Fluorescent In situ

In article <94Dec22.095229est.18900 at gpu.utcc.utoronto.ca>,
ilivne at UTCC.UTORONTO.CA ("I. Livne") wrote:
> Dear netters
> Has any one tried the Molecular Probes ELF substrate for detection of
Digoxigenin probes
> in whole mount in situ hybridization? On paper it looks as promising
alternative to the
> NBT substrate and generates clean Fluorescet signal. Is it for real?
>> Livne Bar
>I.Livne at utoronto.ca
We've tried out the ELF substrate to detect several different
digoxygenin-labeled probes hybridized to whole-mount zebrafish embryos. We
find that it is less sensitive than NBT/BCIP and even slightly less
sensitive than Fast Red. In addition, the reaction product forms a
crytalline precipitate, which is not evenly distributed throughout the
cell, thus reducing cellular resolution. Finally, the substrate comes out
of solution after about 2 hr and sticks to the tissue, causing additional
problems.
Despite the problems, I can see it having some utility. For example, if
you want to mark a domain (as opposed to individual cells) with a
fluorescent signal, this may do it for you.
Hope this helps.
Bruce Appel
--
Bruce Appel
University of Oregon
appel at uoneuro.uoregon.edu